Rapid site-specific DNA inversion in Escherichia coli mutants lacking the histonelike protein H-NS

Abstract

Escherichia coli pilG mutants are thought to have a dramatically higher DNA inversion rate as measured by the site-specific DNA inversion of the type 1 pili piUl promoter. DNA sequence of the pilG gene confirmed its identity to the gene encoding the bacterial histonelike protein H-NS. Unlike other histonelike protein complexes that enhance site-specific DNA recombination, the H-NS protein inhibited this process. This inhibition was indicated by the increased inversion rate of the pilA promoter region effected by two different mutant pilG alleles. One of these alleles, pilGI, conferred a mutant phenotype only at low temperature attributable to a Tto-G transversion in the -35 sequence of the pilG promoter. The other allele, pilG2-tetR, was an insertion mutation in the pilG coding region that conferred the mutant phenotype independent of temperature. We measured an approximately 100-fold-increased piUl promoter inversion rate in the mutant by exploiting the temperature-dependent expression of pilGI and using a novel rapid-population-sampling method. Contrary to one current view on how the H-NS protein might act to increase DNA inversion rate, we found no evidence to support the hypothesis that DNA supercoiling affected piUA promoter inversion.A number of site-specific recombination systems in procaryotes have been described in which the inversion of a DNA segment is responsible for alternating expression of one or more genes. The classic example of this type of gene regulation is the Hin-mediated H-segment inversion in Salmonella typhimurium dictating the antigenic type of flagellin expressed (42). Other Hin-related inversion systems have been described, including Gin-mediated G-loop inversion in Mu (18) and Cin-mediated C-loop inversion in bacteriophage P1(20). The DNA inversion in these two systems determines expression of alternate tail fiber antigens, thereby dictating the host range of these phage. The Hin, Gin, and Cin invertases that promote the site-specific DNA recombination in these systems are functionally similar. Each of these invertases will effectively promote the inversion of the H, G, and C segments under appropriate conditions (20,23,28).Type 1 pilus expression in Escherichia coli is regulated by the inversion of a 314-bp DNA segment containing the promoter for pilA (fimA), the gene encoding the type 1 pilin monomer (1,24,43). When the pilA promoter segment is in one orientation (ON), pilA is transcribed and the E. coli cells are type 1 piliated. With the promoter in the other orientation (OFF), there is no pilA transcription and the cells are nonpiliated. Though the mechanism of pilA promoter inversion is not completely understood, it is clear that this process is significantly different from the Hin-related inversion systems. The 9-bp inverted repeats flanking the invertible pilA promoter segment share no homology with the sequences that bind the Hin-related invertases (24). Also, the Hin protein does not promote pilA promoter inversion (12). The fimB and pilH (fimE, hyp) genes encode proteins r...

A site-specific recombinase is required for competitive root colonization by Pseudomonas fluorescens WCS365

“…Three global regulators have been described to affect switching induced by fimB and fimE, namely IHF, the histone-like protein H-NS, and the leucine-responsive element (Lrp) (37,(42)(43)(44)(45)(46). Lrp differentially regulates switching in various media and is involved in the stimulation of switching upon addition of aliphatic amino acids (47).…”

The leucine-responsive regulatory protein binds to the fim switch to control phase variation of type 1 fimbrial expression in Escherichia coli K-12

“…Although inversion of thefim switch is considered to be controlled by the products of fimB, fimE, himA, himD, and hns (7,8,13,(14)(15)(16), little is known about the mechanism of the site-specific inversion and its potential regulation. We reported previously that rather than being slow and random, the fim switch is capable of high inversion frequencies (>0.7 per cell per generation) and is regulated by environmental conditions, including temperature and the amino acids alanine, leucine, isoleucine, and valine (11).…”

H‐NS: a modulator of environmentally regulated gene expression

“…However, it was found later that mutants carrying insertions in codon 37 and completely lacking H-NS immunocross-reacting material (e.g. hns-2 and hns206; Kano et al, 1993;Dersch et al, 1994) had no effect on plasmid supercoiling (Kano et al, 1993;Kawula and Orndorff, 1991). As mutants of all three classes had similar effects on expression of proU and bgl, the results strongly suggest that there is no correlation between the effect of hns mutants on overall supercoiling and the effect on gene-expression phenotypes.…”